The present application claims priority to German Application No. DE 10045496.8, which was filed on Sep. 13, 2000, the entire contents of which is incorporated herein by reference.
1. Field of the Invention
The present invention provides nucleotide sequences from Coryneform bacteria which code for the PtsI protein and a process for the fermentative preparation of amino acids using bacteria in which the ptsI gene is enhanced.
2. Discussion of the Background
L-Amino acids, in particular L-lysine, are used in human medicine, in the pharmaceuticals industry and in the foodstuffs industry, particularly in animal nutrition.
It is known that amino acids are prepared by fermentation from strains of Coryneform bacteria, in particular Corynebacterium glutamicum. Because of their great importance, work is constantly being undertaken to improve amino acid preparations. Improvements can relate to the fermentation means, such as stirring and supply of oxygen; the composition of the nutrient media, such as the sugar concentration during the fermentation; working up the product form, for example, using ion exchange chromatography; or by altering the intrinsic output properties of the microorganism itself.
Methods of mutagenesis and mutant selection are used to improve the output properties of microorganisms. Strains that are resistant to antimetabolites or are auxotrophic for metabolites of regulatory importance and produce amino acids may be obtained in this manner.
Methods of the recombinant DNA technique have also been employed for some years to improve Corynebacterium strains which produce L-amino acid, by amplifying individual amino acid biosynthesis genes and investigating the effect on the amino acid production.
However, there remains a critical need for improved methods of producing L-amino acids and thus for the provision of strains of bacteria producing higher amounts of L-amino acids. On a commercial or industrial scale even small improvements in the yield of L-amino acids, or the efficiency of their production, are economically significant. Prior to the present invention, it was not recognized that enhancing the ptsI gene encoding the enzyme phosphotransferase system enzyme I (PtsI) would improve L-amino acid yields.
One object of the present invention is providing a new process adjuvant for improving the fermentative production of L-amino acids, particularly L-lysine and L-glutamate. Such process adjuvants include enhanced bacteria, preferably enhanced Coryneform bacteria which express enhanced levels of phosphotransferase system enzyme I which is encoded by the ptsI gene.
Thus, another object of the present invention is providing such a bacterium, which expresses enhanced amounts of phosphotransferase system enzyme I or gene products of the ptsI gene.
Another object of the present invention is providing a bacterium, preferably a Coryneform bacterium, which expresses a polypeptide that has enhanced phosphotransferase system enzyme I activity.
Another object of the invention is to provide a nucleotide sequence encoding a polypeptide which has a phosphotransferase system enzyme I sequence. One embodiment of such a sequence is the nucleotide sequence of SEQ ID NO: 1. Other embodiments of the sequence are the nucleotide sequences of SEQ ID NOS:3 and 6.
A further object of the invention is a method of making a phosphotransferase system enzyme I or an isolated polypeptide having phosphotransferase system enzyme I activity, as well as use of such isolated polypeptides in the production of amino acids. One embodiment of such a polypeptide is the polypeptide having the amino acid sequence of SEQ ID NO: 2. Another embodiment of the sequence is the amino acid sequence of SEQ ID NO:4.
In one embodiment the invention provides isolated polypeptides comprising the amino acid sequences in SEQ ID NOS: 2 and 4.
Other objects of the invention include methods of detecting nucleic acid sequences homologous to SEQ ID NO: 1, particularly nucleic acid sequences encoding polypeptides that have the phosphotransferase system enzyme I activity, and methods of making nucleic acids encoding such polypeptides.
The above objects highlight certain aspects of the invention. Additional objects, aspects and embodiments of the invention are found in the following detailed description of the invention.